A dominant role for the Raf-MEK pathway in forskolin, 12-O-tetradecanoyl-phorbol acetate, and platelet-derived growth factor-induced CREB (cAMP-responsive element-binding protein) activation, uncoupled from serine 133 phosphorylation in NIH 3T3 cells

In this study we describe that platelet-derived growth factor (PDGF), 12-O- tetradecanoyl-phorbol-acetate (TPA), and forskolin induced CREB (cAMP-respo nsive element-binding protein) Ser-133 phosphorylation with comparable magn itude and kinetics in NIH 3T3 cells. While forskolin was the most potent ac tivator of CREB, TPA or PDGF modestly increased CREB activity. The role of protein kinase C, protein kinase A, and the Raf-MEK kinase pathway in the a ctivation and Ser-133 phosphorylation of CREB by these three stimuli was in vestigated. We found that inhibition of the Raf-MEK kinase pathway efficien tly blocks transcriptional activation of CREB by all three stimuli. This do minant involvement of Raf-MEK in CREB transcriptional activation seems to b e uncoupled from CREB Ser-133 phosphorylation. We further demonstrate that although inhibition of Raf-MEK represses forskolin-induced CREB activation, forskolin by itself failed to activate ERK1/2 and Elk-1 mediated transcrip tion. These results suggest that a basal level of Raf-MEK activity is neces sary for both PDGF- and forskolin-induced CREB activation, independent of C REB Ser-133 phosphorylation.